The in vitro uptake of albumin by adult Haemonchus contortus is altered by extracorporeal digestion.
FITC (fluorescein isothiocyanate)-labeled albumin was used as a substrate to further characterize the digestion of host blood proteins by adult Haemonchus contortus. Isolated H. contortus intestine degraded FITC-albumin; degradation was completely inhibited by E-64, a specific inhibitor of cysteine proteases. The in vitro uptake and degradation of FITC-albumin by parasites was also evaluated. Uptake of fluorescence was demonstrated; greater than 50% of the fluorescence was associated with degraded FITC-albumin. However, both the uptake and degradation of FITC-albumin were reduced by about 85%. Additionally, when parasites were incubated in the presence of specific inhibitors of cysteine proteases, degradation of FITC-albumin was shown to occur rapidly in the media during in vitro incubation. This degradation was blocked completely by the presence of cysteine protease inhibitors. These results indicate that the rapid extracorporeal digestion of FITC-albumin (mediated by secreted cysteine proteases) produce relatively high concentrations of low molecular weight FITC-labeled fragments and alters the kinetics of uptake of fluorescence by the parasite. These fragments rather than FITC-albumin are rapidly taken up by the parasites. Thus in vitro demonstration of a definitive role for the cysteine proteases as intestinal digestive enzymes in H. contortus is compromised by the enzyme's extracorporeal activity. This extracorporeal cysteine protease activity may represent the enzyme's predominant activity.
Fetterer
R H
RH
Parasite Biology and Epidemiology Laboratory, Beltsville Agricultural Research Center, MD 20705, USA. rfettere@ggpl.arsusda.gov
Rhoads
M L
ML
D000018:Abomasum / Q000469:parasitology*; D000818:Animals; D001692:Biological Transport; D001711:Biotransformation; D004063:Digestion; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D006189:Haemonchus / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D007814:Larva; D012235:Rhodamines / Q000493:pharmacokinetics; D012710:Serum Albumin, Bovine / Q000493:pharmacokinetics*; D012756:Sheep
D012235:Rhodamines; C039217:fluorescein isothiocyanate bovine serum albumin; D012710:Serum Albumin, Bovine; D016650:Fluorescein-5-isothiocyanate; C029773:rhodamine B